Diabetes is characterized by impaired glucose metabolism manifesting itself among other things by an elevated blood glucose level in the diabetic patient. Underlying defects lead to a classification of diabetes into two major groups: type I diabetes, or insulin dependent diabetes mellitus (IDDM), which arises when patients lack beta-cells producing insulin in their pancreatic glands, and type 2 diabetes, or non-insulin dependent diabetes mellitus (NIDDM), which occurs in patients with an impaired beta-cell function and alterations in insulin action.
Type I diabetic patients are currently treated with insulin, while the majority of type 2 diabetic patients are treated with agents that stimulate beta-cell function or with agents that enhance the tissue sensitivity of the patients towards insulin. Over time almost one-half of type 2 diabetic subjects lose their response to these agents and then must be placed on insulin therapy. The drugs presently used to treat type 2 Diabetes include:
Alpha-glucosidase inhibitors (PRECOSE®, VOGLIBOSE™, and MIGLITOL®). Alpha-glucosidase inhibitors reduce the excursion of postprandial glucose by delaying the absorption of glucose from the gut. These drugs are safe and provide treatment for mild to moderately affected diabetic subjects. However, gastrointestinal side effects have been reported in the literature.
Insulin sensitizers. Insulin sensitizers are drugs that enhance the body's response to insulin. Thiozolidinediones such as REZULIN™ (troglitazone) activate the PPAR gamma receptor and modulate the activity of a set of genes that have not been well described. Although effective, these drugs have been associated with liver toxicity. Because of hepatotoxicity, REZULIN has been withdrawn from the market.
Insulin secretagogues (sulfonylureas and other agents that act by the ATP-dependent K+ channel). SFUs are standard therapy for type 2 diabetics that have mild to moderate fasting glycemia. The SFUs have limitations that include a potential for inducing hypoglycemia, weight gain, and high primary and secondary failure rates. 10 to 20% of initially treated patients fail to show a significant treatment effect (primary failure). Secondary failure is demonstrated by an additional 20-30% loss of treatment effect after six months on an SFU. Insulin treatment is required in 50% of the SFU responders after 5-7 years of therapy (Scheen, A. J., et al., Diabetes Res. Clin. Pract. 6:533-543 (1989)).
GLUCOPHAGE™ (mefformin HCl) is a biguanide that lowers blood glucose by decreasing hepatic glucose output and increasing peripheral glucose uptake and utilization. The drug is effective at lowering blood glucose in mildly and moderately affected subjects and does not have the side effects of weight gain or the potential to induce hypoglycemia. However, GLUCOPHAGE has a number of side effects including gastrointestinal disturbances and lactic acidosis. GLUCOPHAGE is contraindicated in diabetics over the age of 70 and in subjects with impairment in renal or liver function. Finally, GLUCOPHAGE has the same primary and secondary failure rates as the SFUs.
Insulin treatment is instituted after diet, exercise, and oral medications have failed to adequately control blood glucose. This treatment has the drawbacks that it is an injectable, that it can produce hypoglycemia, and that it causes weight gain.
Because of the problems with current treatments, new therapies to treat type 2 diabetes are needed. In particular, new treatments to retain normal (glucose-dependent) insulin secretion are needed. Such new drugs should have the following characteristics: dependent on glucose for promoting insulin secretion, i.e. produce insulin secretion only in the presence of elevated blood glucose; low primary and secondary failure rates; and preserve islet cell function. The strategy to develop the new therapy disclosed herein is based on the cyclic adenosine monophosphate (cAMP) signaling mechanism and its effects on insulin secretion.
Cyclic AMP is a major regulator of the insulin secretion process. Elevation of this signaling molecule promotes the closure of the K+ channels following the activation of protein kinase A pathway. Closure of the K+ channels causes cell depolarization and subsequent opening of Ca++ channels, which in turn leads to exocytosis of insulin granules. Little if any effects on insulin secretion occurs in the absence of low glucose concentrations (Weinhaus, A., et al., Diabetes 47: 1426-1435 (1998)). Secretagogues like pituitary adenylate cyclase activating peptide (“PACAP”) and GLP-1 use the cAMP system to regulate insulin secretion in a glucose-dependent fashion (Komatsu, M., et al., Diabetes 46: 1928-1938, (1997)). Insulin secretagogues working through the elevation of cAMP such as GLP-1 and PACAP is also able to enhance insulin synthesis in addition to insulin release (Skoglund, G. et al., Diabetes 49: 1156-1164, (2000). Borboni, P. et al., Endocrinology 140: 5530-5537, (1999)).
PACAP is a potent stimulator of glucose-dependent insulin secretion from pancreatic beta-cells. Three different PACAP receptor types (R1, R2, and R3) have been described (Harmar, A. et al., Pharmacol. Reviews 50: 265-270 (1998)). PACAP displays no receptor selectivities, having comparable activities and potencies at all three receptors. R1 is located predominately in the CNS, whereas R2 and R3 are more widely distributed. R2 is located in the CNS as well as in liver, lungs and intestine. R3 is located in the CNS, pancreas, skeletal muscle, heart, kidney, adipose tissue, testis and stomach. Recent work argues that R3 is responsible for the insulin secretion from beta cells (Inagaki, N. et al., PNAS 91: 2679-2683, (1994)). This insulinotropic action of PACAP is mediated by the GTP binding protein Gs. Accumulation of intracellular cAMP in turn activates the nonselective cation channels in beta cells increasing [Ca++], and promotes exocytosis of insulin-containing secretory granules.
PACAP is the newest member of the superfamily of metabolic, neuroendocrine and neurotransmitter peptide hormones that exert their action through the cAMP-mediated signal transduction pathway (Arimura, Regul. Peptides 37:287-303 (1992)). The biologically active peptides are released from the biosynthetic precursor in two molecular forms, either as a 38-amino acid peptide (PACAP-38) and/or as a 27-amino acid peptide (PACAP-27) with an amidated carboxyl termini (Arimura, supra).
The highest concentrations of the two forms of the peptide are found in the brain and testis (reviewed in Arimura, supra). The shorter form of the peptide, PACAP-27, shows 68% structural homology to vasoactive intestinal polypeptide (VIP). However, the distribution of PACAP and VIP in the central nervous system suggests that these structurally related peptides have distinct neurotransmitter functions (Koves et al., Neuroendocrinology 54:159-169, (1991)).
Recent studies have demonstrated diverse biological effects of PACAP-38, from a role in reproduction (McArdle, Endocrinology 135:815-817 (1994)) to ability to stimulate insulin secretion (Yada et al., J. Biol. Chem. 269:1290-1293 (1994)).
Vasoactive intestinal peptide (VIP) is a 28 amino acid peptide that was first isolated from hog upper small intestine (Said and Mutt, Science 169: 1217-1218, 1970; U.S. Pat. No. 3,879,371). This peptide belongs to a family of structurally-related, small polypeptides that includes helodermin, secretin, the somatostatins, and glucagon. The biological effects of VIP are mediated by the activation of membrane-bound receptor proteins that are coupled to the intracellular cAMP signaling system. These receptors were originally known as VIP-R1 and VIP-R2, however, they were later found to be the same receptors as PACAP-R2 and PACAP-R3. VIP displays comparable activities and potencies at PACAP-R2 and PACAP-R3.
To improve the stability of VIP in human lung fluid, Bolin et al (Biopolymers 37: 57-66, (1995)) made a series of VIP variants designed to enhance the helical propensity of this peptide and reduce proteolytic degradation. Substitutions were focused on positions 8, 12, 17, and 25-28, which were implicated to be unimportant for receptor binding. Moreover, the “GGT” sequence was tagged onto the C-terminus of VIP muteins with the hope of more effectively capping the helix. Finally, to further stabilize the helix, several cyclic variants were synthesized (U.S. Pat. No. 5,677,419). Although these efforts were not directed toward receptor selectivity, they yielded two analogs (designated herein as R3P0 and R3P4) that have greater than 100-fold PACAP-R3 selectivity (Gourlet et al., Peptides 18: 403-408, (1997); Xia et al., J. Pharmacol. Exp. Ther., 281: 629-633, (1997)).
GLP-1 is released from the intestinal L-cell after a meal and functions as an incretin hormone (i.e. it potentiates glucose-induced insulin release from the pancreatic beta-cell). It is a 37-amino acid peptide that is differentially expressed by the Glucagon gene, depending upon tissue type. The clinical data that support the beneficial effect of raising cAMP levels in β-cells have been collected with GLP-1. Infusions of GLP-1 in poorly controlled type 2 diabetics normalized their fasting blood glucose levels (Gutniak, M., et al., New Eng. J. Med. 326:1316-1322, (1992)) and with longer infusions improved the beta cell function to those of normal subjects (Rachman, J. et al., Diabetes 45: 1524-1530, (1996)). A recent report has shown that GLP-1 improves the β-cells' ability to respond to glucose in subjects with impaired glucose tolerance (Byrne M., et al., Diabetes 47: 1259-1265 (1998)). All of these effects, however, are short-lived because of the short half-life of the peptide. Recently Novo Nordisk has discontinued clinical trials with GLP-1. This failure reportedly was due to a very short plasma half-life of the peptide of a few minutes.
EXENDIN 4™. Amylin Pharmaceuticals is conducting Phase I trials with EXENDIN 4 (AC2993), a 39 amino acid peptide originally identified in Gila Monster. Phase II trials have recently begun. Amylin claims preclinical results showing a 4 hour duration of efficacy and efficacy in animal models when AC2993 is administered subcutaneously, orally, and nasally. However, at doses of 0.2 and 0.3 ug/kg, the incidence of headaches, postural hypotension, nausea and vomiting was significant.
There exists a need for an improved peptide that has the glucose-dependent insulin secretagogue activity of PACAP, GLP-1, or EXENDIN 4, and yet has fewer side-effects.